Damage effect of low frequency and low intensity ultrasound combined with levofloxacin-loaded nanoparticles on Bacille Calmette-Guerin biofilm

Abstract

Objective To explore the damage effect of low frequency and low intensity ultrasound (LFLIU) combined with levofloxacin-loaded PLGA-PEG nanoparticles (LVFX-NPs) on Bacille Calmette-Guerin (BCG) biofilm. Methods Double-emulsification was used to prepare LVFX-NPs, and the physical features of the nanoparticles were detected. XTT assay and crystal violet staining were performed to detect the biofilm activity and biomass changes after treatment of ultrasound combined with LVFX-NPs. SYTO9/PI staining was carried out to qualitatively observe the biofilm damage. Flow cytometry was employed to detect the production of reactive oxygen species (ROS) in BCG, and laser confocal microscopy was adopted to observe the permeability changes of the biofilm. Results LVFX-NPs were successfully prepared, in spherical shape and uniform size, good dispersion, particle size of 254.90±9.78 nm, potential -17.83±0.74 mV, and with drug loading content and encapsulation efficiency of (3.50±0.51)% and (67.00±2.69)%, respectively. The biofilm activity was statistically lower in the ultrasound combined with LVFX-NPs group than the control group (P<0.05). Flow cytometry results showed that ultrasound combined with LVFX-NPs could produce a large amount of ROS. The results of laser confocal microscopy indicated that ultrasound could promote nanoparticles to enter the biofilm. Conclusion LVFX-NPs are successfully prepared, and can penetrate into the BCG biofilm under the action of ultrasound and produce ROS to achieve effective sterilization.

Keywords

• low-frequency and low-intensity ultrasound
• bcg biofilm
• nanoparticles
• reactive oxygen species
• synergistic therapy